Antihemorrhagic Activity Research Articles

Pollen morphological studies on selected species of Jatropha L. through light microscopy from Hyderabad, Telangana State.

Pollen morphology is one of the important tools for taxonomic studies and helps in the identification of medicinal plants. Pollen characterization of four species of Jatropha viz., Jatropha gossypifolia, Jatropha integerrima, Jatropha multifida, and Jatropha podagrica. collected from Greater Hyderabad of Telangana State, India during 2022-23 was carried out by using Light Microscope (LM). The pollen types looked similar but varied in size and shape. The pollen grains are spheroidal to circular, isopolar and radially symmetrical. Grains are inapertutate/omniaperturate with surface showing crotonoid pattern. The plants studied in the present work were known for their medicinal use having anti-inflammatory, antidiarrheal, antiophidian, analgesic, antipyretic, antimicrobial, healing, antianemic, antidiabetic, and antihemorrhagic activities and most important in traditional system of medicine.

Ethamsylate Attenuates Mutilated Secondary Pathogenesis and Exhibits a Neuroprotective Role in Experimental Model of Spinal Cord Injury

Deficits in the neuronal connection that succumbs to the impairment of sensory and motor neurons are the hallmarks of spinal cord injury (SCI). Secondary pathogenesis, which initiates after the primary mechanical insult to the spinal cord, depicts a pivotal role in producing inflammation, lesion formation and ultimately causes fibrotic scar formation in the chronic period. This fibrotic scar formed acts as a major hindrance in facilitating axonal regeneration and is one of the root causes of motor impairment. Cascade of secondary events in SCI begins with injury-induced blood spinal cord barrier rupture that promotes increased migration of neutrophils, macrophages, and other inflammatory cells at the injury site to initiate the secondary damages. This phenomenon leads to the release of matrix metalloproteinase, cytokines and chemokines, reactive oxygen species, and other proteolytic enzymes at the lesion site. These factors assist in the activation of the TGF-β1 signaling pathway, which further leads to excessive proliferation of perivascular fibroblast, followed by deposition of collagen and fibronectin matrix, which are the main components of the fibrotic scar. Subsequently, this scar formed inhibits the propagation of action potential from one neuron to adjacent neurons. Ethamsylate, an anti-hemorrhagic drug, has the potential to maintain early hemostasis as well as restore capillary resistance. Therefore, we hypothesized that ethamsylate, by virtue of its anti-hemorrhagic activity, reduces hemorrhagic ischemia-induced neuronal apoptosis, maintains the blood spinal cord barrier integrity, and decreases secondary damage severity, thereby reduce the extent of fibrotic scar formation, and demonstrates a neuroprotective role in SCI.

Davilla elliptica (Dilleniaceae) a. St.-Hil. Ethnomedicinal, phytochemical, and pharmacological aspects: A review

Abstract Davilla elliptica (Dilleniaceae) is a shrub that occurs naturally in the Brazilian Savannah (Cerrado). This shrub has been widely used in Brazilian folk medicine in tea preparing; infusions; and decoctions from leaves and roots for gastritis, ulcers, and diarrhea treatment. Given the therapeutic potential of this species, the present study provides a comprehensive review on its ethnopharmacological uses, botanical aspects, phytochemistry characterization, and in vivo and in vitro pharmacological studies. The relevant studies on D. elliptica were retrieved from the following scientific databases: Web of Science, Google Scholar, Scopus, SciFinder, PubMed, Scielo, and ScienceDirect. Additional information derived from other literature sources, such as books and Ph.D. and MSc theses, was also considered after a critical review. Ethnopharmacological reports of D. elliptica revealed the use of infusions from roots with tonic and purgative effects, whereas leaves have been used as anti-inflammatory and anti-ulcerative treatments, as well as to treat diarrhea and gastric disorders. Furthermore, fresh leaves have been used in baths to treat swelling. Phytochemical studies have led to the isolation of several compounds, including flavonoids derived from quercetin and myricetin, and tannins, which are known to exhibit relevant pharmacological activities. In vitro assays using crude extracts and/or isolated fractions and chemical constituents from D. elliptica have evidenced antibacterial, cytotoxic activities, while in vivo studies have proven gastroprotective, anti-inflammatory, antihemorrhagic, and antibacterial activities, corroborating ethnomedicinal reports. No relevant side effects has been reported previously, and some studies found a low toxicity and genotoxicity. However, further studies are still needed to ensure the safety of its therapeutic use. Future studies should focused on molecular mechanisms involved in bioactive components’ effects based on both fractions and isolated compounds, creating great opportunities for new investigations.

Natural Inhibitors of Snake Venom Metalloproteinases

Snakebites are a hazard in the tropical world. Although antivenom therapy is effective, it is beset with inherent drawbacks. A better understanding of the major components of snake venoms and their neutralisation will help in improving snakebite treatment. Snake venom metalloproteinases (SVMPs) are responsible for severe haemorrhage, the inhibition of coagulation and platelet aggregation, observed in the victims of snakebite envenoming. Inhibitors from various sources including medicinal plants, animal venoms, and sera are sought to block the pharmacological functions of SVMPs. In this review, we describe the interaction of natural inhibitors with SVMPs. To understand their inhibitory mechanisms, we focussed on the complex structures of these inhibitors and SVMPs. There are three distinct classes of inhibitors; namely, chelators, competitive inhibitors, and non-competitive inhibitors. A small number of inhibitors show their anti-hemorrhagic activity in invivo animal models in treatment mode, but most studies evaluate either invitro neutralisation of enzymatic activity or invivo effects in pre-incubation protocols. We propose the distinct strategies and limitations to design either broad-spectrum or highly selective SVMP inhibitors. The goal of designing broad-spectrum inhibitors against SVMPs capable of effective treatment of snakebites without toxicity has been elusive, probably because of the narrow molecular footprint of inhibitors against a large number of SVMPs with distinct molecular surfaces. Our ability to design highly selective inhibitors is limited by the lack of information of interactions between selective inhibitors and SVMPs. Comparisons of structures of hemorrhagic and non-hemorrhagic SVMPs revealed different distributions of electric charge on the surface of SVMPs, which may be exploited to design specific inhibitors. The specific inhibitors may also be useful to identify target molecules of the SVMPs and help to understand their mechanism of action.

Skład chemiczny i właściwości biologiczne rożnika przerośniętego (Silphium perfoliatum L.)

A cup plant (Silphium perfoliatum L.) is a poorly-known crop in the country. It came to Europe from North America in the 18th century. It can be propagated by dividing rhizomes or by sowing seeds. This plant is undemanding in terms of alimentary requirements and resistant to cold and pests. Due to the high content of nutrients and the yield of green mass a cup plant is used for fodder purposes. A cup plant contains numerous secondary metabolites, such as phenolic acids, flavonoids, terpenes, essential oil, tannins, vitamins and mineral components. It is characterized by antioxidant, antibacterial, antifungal, cytostatic and immunosuppressive activity. Research on laboratory animals indicates that the cup plant accelerates wound healing and indicate hypolipemic and estrogenic activity. Numerous reports confirm, that this plant also has a strengthening, diuretic, anti-inflammatory, antihemorrhagic and spasmolytic activity. In addition, it is used in the healing diseases of the liver and spleen, and also heals gastric and duodenal ulcers.

Cirsium Japonicum DC ingredients-loaded silk fibroin nanofibrous matrices with excellent hemostatic activity

Regenerated Silk fibroin (SF) from Bombyxmori has emerged in recent years as candidates for wound dressing due to its excellent biocompatibility and low inflammatory response. Cirsium Japonicum DC (CJDC) is a kind of Chinese herb medicine with evident hemostatic and anti-inflammatory analgesic effect. In this study, we aimed to manufacture a novel green electrospun SF nanofibrous matrice loading ingredients extracted from CJDC which should be useful in hemostasis as well as wound healing. FTIR spectra showed that the CJDC extracts were successfully loaded into the SF nanofibrous matrices, which appeared morphology of porous mesh structure. The as-prepared matrices inherited the good skin affinity and hemocompatibility of SF, as indicated by the skin cell viability assay and hemolysis test in vitro. Furthermore, according to the results of plasma recalcification profiles test, the incorporation of CJDC extracts significantly improved the hemostatic performance of SF nanofibrous matrices. Interestingly, the SF nanofibrous matrices loading CJDC components extracted by petroleum ether (peE-CJDC@SF) showed the best antihemorrhagic activity among the tested samples (with astringent time of 9.5 min), even better than some clinically used products such as absorbable gelatin sponge (AGS) and aseptic hemostyptic gauze (AHG). Our findings suggested the peE-CJDC@SF nanofibrous matrices could be a promising candidate using as hemostatic material and wound dressing.

Isolation and characterization of a novel metalloprotease inhibitor from Bothrops alternatus snake serum

Resistance of snakes and some other animals to snake envenomation has been attributed to soluble factors present in their tissues. Here we report the isolation of a novel metalloprotease inhibitor from Bothrops alternatus snake serum (named BaltMPI) with high purity, using a four-step chromatographic method. BaltMPI has molecular weights of 60.5 and 42.4kDa, as determined by SDS-PAGE and mass spectrometry, respectively, and pI=5.27. The first 60 amino acids from the N-terminal region of BaltMPI, determined by Edman’s degradation, showed high homology (97%) with the snake venom metalloprotease inhibitor (SVMPI) BJ46a and other SVMPIs (78–82%). The chromatographic fractions and purified BaltMPI exhibited anti-hemorrhagic activity against Batroxase and BjussuMP-I. BaltMPI was stable over wide ranges of pH (1, 5, 8, and 9) and temperature (−80, −20, 4, 60, and 100°C), and suppressed the fibrinogenolytic, fibrinolytic, and azocaseinolytic activities of Batroxase. BaltMPI specifically inhibited the activity of metalloproteases, without affecting the activity of serine proteases. Together, our results suggest that BaltMPI and other SVMPIs are promising molecules for the treatment of snake envenomation, in particular that caused by Bothrops sp.

Improving Dissolution and Photostability of Vitamin K3 via Cocrystallization with Naphthoic Acids and Sulfamerazine

Menadione (MD), also known as vitamin K3, has been widely applied in fortified food, feed, and the nutrition industry for its antihemorrhagic activity. However, the poor photostability in the solid state has greatly affected its biological performance and limited its applications. With the objective to alter the topochemical photoreactivity, a cocrystallization approach was employed, and three MD cocrystals with naphthoic acids and sulfamerazine were designed and prepared. Single crystal structures were determined, and solid-state characterization were performed. Solid-state UV–vis spectra revealed a significant red-shift of UV absorption in the cocrystal solids resulting in variant color differences. These physicochemical changes may be attributed to the enhanced π···π interactions and change-transfer interactions within these molecular complexes. More importantly, the newly synthesized cocrystals displayed better dissolution behavior and superior photostability with respect to MD itself. No obvious degr...

Anti-hemorrhagic effect of hydro-alcoholic extract of the leaves of Mikania glomerata in lesions induced by Bothrops jararaca venom in rats

To investigate the effects of hydroalcoholic leaf extract of Mikania glomerata Spreng (Asteraceae) on the activity of Bothrops jararaca snake venom in Wistar rats. Fifty four rats Wistar were divided into six groups of nine animals in each: control treated with saline; control treated with B. jararaca venom; control treated with M. glomerata extract; B. jararaca venom incubated with M. glomerata extract at proportions of 1:1, 1:2, and 1:4. Histopathological and morphometric analysis showed that intradermal administration of snake venom incubated with the hydroalcoholic extract at proportions of 1:1, 1:2 and 1:4 promoted a significant reduction in the number of inflammatory cells and a marked decrease in edema after the third hour. There was also a significant reduction in the intensity of the hemorrhagic halo in animals receiving the snake venom incubated with the extract, with the observation of a progressive and parallel inhibition with increasing proportion of M. glomerata. The Mikania glomerata hydroalcoholic extract exerted effective anti-inflammatory and antihemorrhagic activity against the effects induced by Bothrops jararaca snake venom.

Efficacy of a new colloidal form of low-sialylated polylactide-based erythropoietin in experimental hemorrhagic stroke in rats

The aim of the present work was to develop methods for the preparation of a nanosomal form of low-sialylated recombinant human erythropoietin (ls-rhEPO), which has neurotropic antihemorrhagic activity. Measurements were made of the sorption of erythropoietin to nanoparticles made of lactic acid homopolymer or lactic acid/glycolic acid copolymer, mean nanoparticle size, and the resistance of the resulting complex to aggregation. The most effective sorption (greater than 80%) was achieved using a copolymer with equal proportions of lactic and glycolic acids (50:50). Experiments using an intracerebral post-traumatic hematoma model in rats demonstrated that nanosomal ls-rhEPO had neuroprotective actions, based on its ability to decrease death in the experimental animals (77.8% survived, which was 40% more than in the control group). Nanosomal ls-rhEPO was found to have no effect on hematopoiesis in conditions of chronic administration.

Benzenepolycarboxylic acids with potential anti-hemorrhagic properties and structure–activity relationships

Previously, we reported the structural requirements of the cinnamic acid relatives for inhibition of snake venom hemorrhagic action. In the present study, we examined the effect of benzenepolycarboxylic acids and substituted benzoic acids against Protobothropsflavoviridis venom-induced hemorrhage. Pyromellitic acid (1,2,4,5-benzenetetracarboxylic acid) was found to be a potent inhibitor of hemorrhage, with an IC50 value of 0.035μM. In addition, most of the antihemorrhagic activity of compounds tested in this experiment showed good correlation to acidity.

Contribution of cinnamic acid analogues in rosmarinic acid to inhibition of snake venom induced hemorrhage

In our previous paper, we reported that rosmarinic acid ( 1) of Argusia argentea could neutralize snake venom induced hemorrhagic action. Rosmarinic acid ( 1) consists of two phenylpropanoids: caffeic acid ( 2) and 3-(3,4-dihydroxyphenyl)lactic acid ( 3). In this study, we investigated the structural requirements necessary for inhibition of snake venom activity through the use of compounds, which are structurally related to rosmarinic acid ( 1). By examining anti-hemorrhagic activity of cinnamic acid analogs against Protobothrops flavoviridis (Habu) venom, it was revealed that the presence of the E-enoic acid moiety (–CH CH–COOH) was critical. Furthermore, among the compound tested, it was concluded that rosmarinic acid ( 1) (IC 50 0.15 μM) was the most potent inhibitor against the venom.

Biological and Pathological Studies of Rosmarinic Acid as an Inhibitor of Hemorrhagic Trimeresurus flavoviridis (habu) Venom

In our previous report, rosmarinic acid (RA) was revealed to be an antidote active compound in Argusia argentea (family: Boraginaceae). The plant is locally used in Okinawa in Japan as an antidote for poisoning from snake venom, Trimeresurus flavoviridis (habu). This article presents mechanistic evidence of RA’s neutralization of the hemorrhagic effects of snake venom. Anti-hemorrhagic activity was assayed by using several kinds of snake venom. Inhibition against fibrinogen hydrolytic and collagen hydrolytic activities of T. flavoviridis venom were examined by SDS-PAGE. A histopathological study was done by microscopy after administration of venom in the presence or absence of RA. RA was found to markedly neutralize venom-induced hemorrhage, fibrinogenolysis, cytotoxicity and digestion of type IV collagen activity. Moreover, RA inhibited both hemorrhage and neutrophil infiltrations caused by T. flavoviridis venom in pathology sections. These results demonstrate that RA inhibited most of the hemorrhage effects of venom. These findings indicate that rosmarinic acid can be expected to provide therapeutic benefits in neutralization of snake venom accompanied by heat stability.

Changes in the content of hyperoside in aerial parts of Pulmonaria mollis during the vegetation period

Extracts taken from the aerial parts of Pulmonaria mollis Wulf. ex Hornem. (Boraginaceae) have anti-anemic activity. Bioactive compounds having anti-hemorrhagic activity play an important role in the phytotherapy of anemia. Such compounds include flavonoids which can be found in P.mollis. The presence of some flavonoids (predominantly hyperoside) in the aerial parts of P.mollis was established by TLC (SiO2). The content of flavonoids was analysed using spectrophotometric methods with AlCl3. The total amount of flavonoids was calculated as the hyperoside equivalent.

Snake fetuin: Isolation and structural analysis of new fetuin family proteins from the sera of venomous snakes

Novel proteins were isolated from the sera of Chinese Mamushi ( Gloydius blomhoffi brevicaudus) and Habu ( Trimeresurus flavoviridis). The primary structures of these proteins were determined by protein sequencing, and the nucleotide sequences were established by cDNA cloning from the liver mRNAs. They belonged to the fetuin family having a double-headed cystatin-like domain and a His-rich domain, akin to HSF, an antihemorrhagic factor isolated from Habu serum. They showed no antihemorrhagic activity and were designated HSF-like proteins (HLPs). Mamushi serum contained two different HLPs termed HLP-A and HLP-B. Both HLP-A and Habu HLP had a unique 17-residue deletion in their His-rich domains. HLP-B comprised two glycosylated polypeptide chains and inhibited the precipitation of calcium phosphate as potently as does bovine fetuin. HLP-B was hence identified as a snake fetuin. The phylogenetic analysis of the fetuin family of proteins showed that antihemorrhagins and HLPs have evolved from this snake fetuin.

Properties and cDNA cloning of antihemorrhagic factors in sera of Chinese and Japanese mamushi ( Gloydius blomhoffi)

An antihemorrhagic protein has been isolated from the serum of Chinese mamushi ( Gloydius blomhoffi brevicaudus) by using a combination of ethanol precipitation and a reverse-phase high-performance liquid chromatography (HPLC) on a C8 column. This protein—designated Chinese mamushi serum factor (cMSF)—suppressed mamushi venom-induced hemorrhage in a dose-dependent manner. It had no effect on trypsin, chymotrypsin, thermolysin, and papain but inhibited the proteinase activities of several snake venom metalloproteinases (SVMPs) including hemorrhagic enzymes isolated from the venoms of mamushi and habu ( Trimeresurus flavoviridis). A similar protein (Japanese MSF, jMSF) with antihemorrhagic activity has also been purified from the sera of Japanese mamushi ( G. blomhoffi). The N-terminal 70 and 51 residues of the intact cMSF and jMSF were directly analyzed; a similarity between the sequences of two MSFs to that of antihemorrhagic protein (HSF) from habu serum was noticed. To obtain the complete amino acid sequences of MSFs, cDNAs encoding these proteins were cloned from the liver mRNA of Chinese and Japanese vipers based on their N-terminal amino acid sequences. The mature forms of both MSFs consisted of 305 amino acids with a 19-residue signal sequence, and a unique 17-residue deletion was detected in their His-rich domains.

Antibacterial, antisecretory and antihemorrhagic activity of Azadirachta indica used to treat cholera and diarrhea in India

Indigenous uses of Azadirachta indica A. juss (Maliaceae) (locally known as neem) leaves in different parts of India for curing gastrointestinal disorder such as diarrhea and cholera is wide spread. The objective of the present study was to evaluate the antibacterial and antisecretory activity of neem extract against Vibrio cholerae, a causative agent of watery diarrhea such as cholera. The methanol extract of neem leaf was tested for its antibacterial, antisecretory and antihemorrhagic activity against Vibrio cholerae. Azadirachta indica extract had significant antibacterial activity against the multi-drug-resistant Vibrio cholerae of serotypes O1, O139 and non-O1, non-O139. The minimum inhibitory concentration reached by 50% (MIC 50) and 90% (MIC 90), and minimum bactericidal concentration for the extract were 2.5, >5, and 10 mg/ml, respectively. Neem extract showed antisecretory activity on Vibrio cholerae induced fluid secretion in mouse intestine with inhibition values of 27.7%, 41.1%, 43.3%, 57.0%, and 77.9% at doses of 100, 200, 300, 450 and 1800 mg/kg, respectively. Oral administration of the extract inhibited hemorrhage induced by Vibrio cholerae in mouse intestine at a dose ≥300 mg/kg. The results obtained in this study give some scientific support to the uses of neem employed by the indigenous people in India employed for the treatment of diarrhea and dreadful disease cholera.

Inhibition of Proteolytic and Hemorrhagic Activities by Ethyl Acetate Extract of Eclipta prostrata. Against Malayan Pit Viper Venom

The ethyl acetate extract of Eclipta prostrata. L. (Asteraceae) was evaluated for its antivenom potential against Calloselasma rhodostoma. Kuhl (Viperidae) (Malayan pit viper; MPV) venom. The partially purified ethyl acetate extract (PEE) was found to contain 47% wedelolactone as its major constituent. PEE and wedelolactone demonstrated strong antiproteolytic and antihemorrhagic activities against MPV venom in a dose-dependent manner. The extract, at 5 mg/mL, could inhibit proteolytic activity of 100 µg of the venom and hemorrhagic activity of 3 minimum hemorrhagic doses (MHD) to 95% and 68%, respectively. At the same concentration, wedelolactone could neutralize the proteolytic activity at around 76% and, at doses of 0.25–1 mg/mL, offered protection against hemorrhagic activity of the venom in the range 3–35%. Both PEE and wedelolactone displayed partial anti–phospholipase A2 activity (21% for PEE and 7% for wedelolactone) and could not neutralize the lethal effect of either 2LD50 or 4LD50 of MPV venom.

Active fragments of the antihemorrhagic protein HSF from serum of habu ( Trimeresurus flavoviridis)

Certain snakes have antihemorrhagic proteins in their sera. Habu serum factor (HSF), an antihemorrhagic protein isolated from the serum of the Japanese habu snake ( Trimeresurus flavoviridis) is composed of two cystatin-like domains (D1 and D2) and a His-rich domain, and it inhibits several snake venom metalloproteinases (SVMPs). The activity of HSF can be abolished by trinitrophenylation of Lys residues with 2,4,6-trinitrobenzene sulphonic acid. Upon complex formation of HSF with SVMP, however, the loss of its inhibitory activity by the chemical modification was suppressed, and Lys 15, Lys 41, and Lys 103 residues in HSF were not trinitrophenylated. In order to identify the domain that is critical to the inhibitory activity on SVMPs, native HSF was digested with papain followed by cleavage with cyanogen bromide, yielding a low-molecular mass fragment that was composed of two peptide chains (residues 5–89 and 312–317) linked by a disulfide bond. This fragment inhibited several SVMPs and showed significant antihemorrhagic activity. This indicates that the N-terminal half of D1 is indispensable for the antihemorrhagic activity of HSF. Furthermore, a three-dimensional model of two cystatin-like domains constructed by the homology modeling has indicated that three Lys residues (15, 41, and 103) are exposed to the same surface of HSF molecule.

Standardization of an antihemorrhagic potency test of antivenoms prepared from two different Agkistrodon halys venoms

To establish Korea National Standards for venoms and antivenoms, it is necessary to have standardized assay methods. In this study, we standardized a method to evaluate the antihemorrhagic potency of two horse-derived antivenoms using rabbit intracutaneous injection. We expressed the capability of these antivenoms to neutralize the hemorrhagic activities triggered by the venoms of Agkistrodon halys from Japan and Jiangzhe Agkistrodon halys from China as Minimum Hemorrhagic Dose (MHD). We also performed cross-neutralization tests employing the parallel line assay on different pairings of venoms and antivenoms to check the possibility of using Jiangzhe Agkistrodon halys venom as a substitute for the standard Agkistrodon halys venom in measurements of the antihemorrhagic activity, since A. halys venom is not easily available. Slope function ratio (S.R.) was 0.957 for Agkistrodon halys venom either with Agkistrodon halys antivenom or with Jiangzhe Agkistrodon halys antivenom (p>0.05). Similarly, S.R. was 0.348 for Jiangzhe Agkistrodon halys venom either with Agkistrodon halys antivenom or with Jiangzhe Agkistrodon halys antivenom (p>0.05). Thus, in this study we established antihemorrhagic potency test methods for both Agkistrodon halys and Jiangzhe Agkistrodon halys antivenoms and we could also show it is possible to use Jiangzhe Agkistrodon halys venom as a standard.